A 2-piece gene therapy for difficult-to-treat diseases

In 2023, the US Food and Drug Administration approved the first gene therapy for Duchenne muscular dystrophy (DMD). But that therapy, Elevidys, can‘t replace the entire dystrophin gene, the absence of which causes DMD. Instead, it inserts a gene that codes for a smaller version of the dystrophin protein because the whole gene is too big to be replaced by one adeno-associated virus (AAV) vector. But researchers theorize that the closer one can get to replicating the entire gene, the better the therapeutic results will be.

Dividing the gene into multiple fragments for delivery by multiple adenovirus vectors could solve the problem. To test this possibility, researchers from the University of Rochester developed a technology they call StitchR that mends two messenger RNA fragments from a split gene together (Science 2024, DOI: 10.1126/science.adp8179). Once the gene fragments are transcribed into mRNA, ribozymes cleave the ends of the mRNA that need to join together, creating ends that the cell machinery stitches together into one strand of mRNA.

The healthy, untruncated dystrophin gene is still too big to fit into two AAVs, so the researchers substituted a smaller version of the gene, called ΔH2-R15. It’s still too big to fit into a single AAV, but it can fit into two. Even though it’s not the full dystrophin gene, the researchers say it’s still better than the even-smaller microdystrophins that can fit into one AAV because it includes more information that’s important for functionality.

In a mouse model of DMD in which the dystrophin gene was knocked out, the researchers found that injecting the StitchR-enabled ΔH2-R15 gene restored the amount of ΔH2-R15 to levels comparable to the dystrophin protein levels of the control mice, and it restored some biomarkers of muscle damage in DMD mice to the same phenotype as in the control mice.

Treating DMD effectively will likely require more research to fit larger versions of the dystrophin gene into the StitchR process. But the technology could be applied to an array of other conditions.

“Right now, we’re building vectors for all these other large-gene diseases that could be treated with gene therapy,” says Doug Anderson, the lead author on the paper from the University of Rochester. His company Scriptr Global is currently developing this platform, and the muscular dystrophy program has been sublicensed to CANbridge Pharmaceuticals, which he says is “very interested in moving that forward towards a therapeutic.”

John Counsell, a professor in medical innovation at University College London who was not involved with the research, says that this technique is a step forward, but logistical challenges need to be addressed. Making one AAV-enabled gene therapy is expensive as it is, and the cost to make two would be even higher. Additionally, not every targeted cell would receive both AAVs, raising the possibility that only one fragment would be translated in a cell that got only one AAV. But he notes that the authors did check for this possibility and found that expression of single fragments was low.

“I think there’s some more steps to go through, but they’re really engineering problems, and they’ve shown the thing that they’ve created here has potentially got promise,” he says.